*After step 8 add 400µl PCA solution to the tube, invert to mix, and centrifuge for 3 minutes at maximum speed. Collect the upper phase by pipetting into a new tube and proceed with step 9. This step helps remove any residual proteins.

+

*After step 9 add 400µl PCA solution to the tube, invert to mix, and centrifuge for 3 minutes at maximum speed. Collect the upper phase by pipetting into a new tube and proceed with step 10. This step helps remove any residual proteins.

*If using gram-positive cells: between steps 3 and 4 add 10mg of lysozyme and incubate for 30 minutes at 37°C.

+

-

===References===

+

===Scale-up===

+

Even higher concentrations of plasmid have been obtained by scaling up this protocol 5x in 15ml centrifuge tubes. However, to do this a larger (i.e. slower) centrifuge was necessary to accomodate the larger tubes. The adjusted centrifugation steps were as follows:</br>

This protocol is based on a protocol described by Sambrook and Russell in their manual "Molecular Cloning" (2001) p:1.32-1.34

This protocol is based on a protocol described by Sambrook and Russell in their manual "Molecular Cloning" (2001) p:1.32-1.34

+

+

==BioCoder version==

+

Following is the Miniprep/GET Buffer protocol in BioCoder, a high-level programming language for expressing biology protocols. What you see here is the auto-generated text ouput of the protocol that was coded up in BioCoder (see Source code). More information about BioCoder can be found on my home page. Feel free to mail me your comments/ suggestions.[[User:Vaishnavi Ananth|Vaishnavi]]

Centrifuge at maximum speed for 3 minutes. Make sure the pellet is toward the outside.

Discard the supernatant and air dry the pellet for 10-15 minutes.

Once the pellet is COMPLETELY DRY resuspend the plasmid DNA in 20 ul of TE or distilled water. The DNA will contain RNA contamination, which can be removed by resuspending in TE with RNAse.

Optional Steps

After step 3 add 10mg lysozyme and incubate for 30 mins before proceeding with step 5. This step is essential for lysing gram-positive cells.

After step 9 add 400µl PCA solution to the tube, invert to mix, and centrifuge for 3 minutes at maximum speed. Collect the upper phase by pipetting into a new tube and proceed with step 10. This step helps remove any residual proteins.

Notes

Scale-up

Even higher concentrations of plasmid have been obtained by scaling up this protocol 5x in 15ml centrifuge tubes. However, to do this a larger (i.e. slower) centrifuge was necessary to accomodate the larger tubes. The adjusted centrifugation steps were as follows:</br>

References

This protocol is based on a protocol described by Sambrook and Russell in their manual "Molecular Cloning" (2001) p:1.32-1.34

BioCoder version

Following is the Miniprep/GET Buffer protocol in BioCoder, a high-level programming language for expressing biology protocols. What you see here is the auto-generated text ouput of the protocol that was coded up in BioCoder (see Source code). More information about BioCoder can be found on my home page. Feel free to mail me your comments/ suggestions.Vaishnavi